This project on protein biosynthesis is divided into two parts: 1) Studies on the dissociation of ribosomes from mRNA at the termination codon of a cistron. We have isolated a pure ribosome releasing (RR) factor which functions to release ribosomes from mRNA at the termination codon. Continuing our preliminary observation which suggests that, in the absence of RR factor, ribosomes will stay on mRNA and continue translation beyond the termination codon, we would like to examine whether the intercistronic polynucleotide sequence is translated. The number of nucleotides in the intercistronic region between the A cistron and the coat cistron of R17 phage and between the A cistron and the B cistron of tryptophan operon are not multiples of three. With the use of these cistrons, we shall determine whether ribosomes would read the distal cistrons "out of phase" in the absence of RR factor. We shall isolate a temperature sensitive mutant having a lesion in the RR factor. The mechanism of action of this factor in releasing ribosomes from mRNA will also be studied. A corresponding factor in eucaryotes will be sought. 2) Studies on protein biosynthesis in ghost infected cells. Continuing our work on the ghost and phage infected cells, we shall study the effect of phage and phage ghost on the synthesis of specific proteins such as the lipoprotein of the envelope, alkaline phosphatase of the periplasm, and the L7 protein of ribosomes. Since ribosomes remain intact in the ghost infected cells, and yet no protein synthesis takes place in these cells, we will explore the possibility that the relationship between membrane and ribosomes may be critical for the maximum in vivo activity of ribosomes. We have recently succeeded in partially restoring the biosynthetic activity of ghost infected cells. This "reconstituted" system is amenable to biochemical manipulations and yet has a morphological cellular integrity. We shall study the synthesis of the membrane lipoprotein, periplasmic alkaline phosphatase, and ribosomal protein and their distribution into outer membrane, periplasm and ribosomes respectively in this "reconstituted" ghost infected cell system. In addition, membrane lipid synthesis will be studied in this system.